P1578: SERUM EXTRACELLULAR VESICLE MIRNA PROFILING TO DETERMINE EXTRACORPOREAL PHOTOPHERESIS TREATMENT RESPONSE IN GRAFT VERSUS HOST DISEASE PATIENTS
Authors: Kimberly Schell, Aisling Flinn, Lucas Cortes, Chong Yun Pang, Andrew Gennery, Matthew Collin, Rachel Crossland
Journal: HemaSphere (2023)

Topic: 31. Transfusion medicine Background: Graft versus Host disease (GvHD) is a life-threatening complication of allogeneic haematopoietic stem cell transplantation (HSCT), where donor material recognises host tissue as foreign, resulting in host tissue destruction. Of those receiving HSCT, 40-70% develop GvHD, leading to post-treatment mortality in 40-60% of patients. Steroids frequently act as first line treatment, however they have significant adverse effects as well as the potential for developing steroid refractoriness. Extracorporeal photopheresis (ECP) therapy is a safe alternative, though the precise mechanism of action is unknown, and there are no confirmed biomarkers to determine ECP response. Aims: Our aim is to investigate the role of serum extracellular vesicles (EVs) during ECP therapy for GvHD. Methods: Serum samples were acquired from n=12 paediatric patients undergoing ECP therapy at the Great North Children’s Hospital, Newcastle upon Tyne, UK, before every weekly treatment cycle. EVs were isolated from 4 sequential timepoints (TP) by precipitation (Total Exosome Isolation Reagent) for n=6 complete (CR) and n=6 partial/non-responders (PR). Particle size and concentration were measured using Nanoparticle Tracking Analysis (NanoSight). Total RNA was extracted, concentrated, quantified, and microRNA expression profiling was performed using NanoString technology (n=800 microRNA). Results: Overall EV modal size was significantly smaller for patients who reached PR compared to CR, and this was most pronounced between TP1 and TP2. There was no observed difference in EV production. Analysing the EV microRNA composition using DESeq2 showed that in principle component analysis (PCA), three groups formed clusters; PR and two separate clusters of mostly CR with some PR. This analysis also showed that treatment cycle was not a driving factor, as there was no treatment cycle-based clustering. There were 8 microRNAs that were significantly differentially expressed in CR compared to PR patients following FDR adjustment; hsa-miR-23a-3p was downregulated and hsa-miR-144-3p, hsa-miR-548ar-3p, hsa-miR-378e, hsa-miR-21-5p, hsa-let-7i-5p, hsa-miR-579-3p, and hsa-miR-1283 were upregulated (p<0.05 and Log2 Fold Change range -1.04 – 24.21). Initial miRNA-Target analysis was performed using MIENTURNET, based on miRTarBase, and a threshold of 3 miRNA-target interactions. This revealed PTEN as the main target of 3 miRNAs, namely hsa-miR-21-5p, hsa-miR-144-3p, and hsa-miR-23a-3p. Other targets include APP, JMY, TGFBR3, and YOD1, several of which are involved in tumour suppression. Further assessment of EV surface markers, serum cytokines, and TCR diversity is ongoing. Summary/Conclusion: Preliminary analysis shows the potential for miRNA biomarkers as an indicator of ECP response, as well as shedding light on the mechanisms of ECP action.Keywords: Graft-versus-host disease (GVHD), Extracorporeal photopheresis